The effects of some features of cell membrane structure on cholesterol-phospholipid interactions were examined for the human erythroctye membrane. Studies were made with erythrocytes in which the density of packing of the phospholipid molecules of the external leaflet of the membrane phospholipid bilayer was reduced by up to 40%, by enzymatic hydrolysis of its phosphatidyl choline, followed by extraction of the hydrolysis products. This treatment had no effect on either the rate of cholesterol exchange between erythrocytes and plasma lipoproteins, or on the equilibrium distribution of cholesterol between erythrocytes and (egg) phosphatidyl choline liposomes (expressed in terms of cholesterol to phospholipid ratio). In another approach, the equilibrium distribution of cholesterol between the cell membrane an liposomes prepared from their whole phospholipid extract was studied. Since in both systems the phospholipid molecules are arranged in a bilayer, and specific interactions between cholesterol and the various phospholipid species are the same, a selectivity in this distribution would reflect cell membrane specific factors such as interaction of either, or both, of cholesterol and phospholipid with membrane protein, or the detailed arrangement of the several phospholipid species in the membrane. The equilibrium distribution coefficient was found to be 1.1-1.2, the cell membrane having a lower affinity for cholesterol than the liposomes. Considered solely on an energetic basis, this lower affinity corresponds to only a small, .06 to .10 kcal. moletothe-1, difference between cholesterol-phospholipid interactions in the two systems. However, the lower affinity of the cell membrane could also be the result of the unavailability of 10-20% of membrane phospholipid to mix with cholesterol.